The lipid bilayer of the cellular membrane insulates the intracellular milieu from exposure to hydrophilic compounds. Unlike lipophilic compounds that can diffuse through cellular membranes, water-soluble compounds usually require specific transport mechanisms to gain access to the intracellular space. The regulation of the traffic of polar compounds in both directions across the cellular membrane is a complex process involving several large families of transport proteins.
Most often in cancer research, drug transport is thought of as a mechanism of cellular drug resistance, as drug efflux pumps such as the products of the MDR1 and MRP genes have been shown to be mechanisms of resistance to lipid-soluble anticancer drugs. However, drug transport is a two-way street, and mechanisms also exist for pumping drugs into cells. For polar, water-soluble anticancer agents, drug uptake, and not drug efflux, is the critical determinant of cellular drug accumulation.
Most cancer chemotherapy employs drugs that are lipid-soluble that can easily penetrate the cell membrane of cancer cells. One advantage of using lipid-soluble drugs is that they easily gain intracellular access to different types of cancer cells, so many cancer cells appear to be initially sensitive to these drugs. The disadvantage is that cancer cells learn to increase the activity of drug efflux pumps in the cell membrane to pump lipid-soluble drugs out of the cell, resulting in drug resistance.
In contrast, potential water-soluble anticancer drugs may not survive the preclinical screening process since there is a great deal of variability in the expression of drug transport genes in different types of cancer cells. Variability in transport gene expression may result in variability in accumulation of polar, water-soluble drugs. One approach to more effectively utilize water-soluble anticancer drugs is to identify which of the dozens of transport genes are actually expressed in tumors.
The importance of carrier-mediated anticancer drug uptake is exemplified in reduced folate carrier (RFC) mediated uptake of methotrexate (MTX). Methotrexate (MTX), a reduced folate analogue, is scavenged and retained in cells by mechanisms designed to secure folates from the environment. The major mechanism of MTX uptake at pharmacologic concentrations is the reduced folate carrier (RFC), an OAT transporter with a Km for MTX between approximately 0.8-26 μM. Decreased RFC activity has been observed in several in vitro models of transport-mediated MTX resistance (Biochem. Pharmacol. 11: 1233-1234, 1960). Once rodent and human genes encoding proteins with RFC activity were isolated, the molecular explanations for decreased RFC activity emerged. RFC1 transfection into the transport-deficient MTXR ZR75 cell line resulted in a 20-fold increase in 6-hour MTX uptake and a concomitant 250-fold increase in sensitivity to MTX relative to control cell clones, showing that the RFC1 gene reconstitutes RFC activity and has a significant impact on MTX cytotoxicity (Moscow, et al., Cancer Res. 55: 3790-3794, 1995).
In different cell lines, MTX transport deficiency has been ascribed either to mutations in the RFC gene or in decreased expression of the RFC gene product. Several studies have demonstrated that RFC1 gene expression is an important determinant of sensitivity to MTX. In in vitro studies, we have found that RFC1 RNA levels correlate with MTX sensitivity in a panel of non-selected cell lines, including breast cancer cell lines (Moscow et al., Int J Cancer. 72: 184-190, 1997).
A plethora of genes with the ability to transport MTX out of the cell have been reported, including MRP1, MRP2, MRP3, MRP4, the organic anion transporters hOAT2 and hOAT3, and the mitoxantrone-resistance protein (BCRP/MXR). However, despite the multitude of MTX export genes, clinical studies have shown a relationship between the expression of RFC1, the mechanism of MTX uptake, and prognosis in Acute Lymphoid Leukemia (ALL) and osteosarcoma. As a result, RFC1 expression and MTX uptake are now implicated as determinants of clinical sensitivity in several types of tumors. Thus, the role of RFC1 in mediating sensitivity of its cytotoxic drug substrates has become a prototype that illustrates the potential role of transporters, like OAT and OCT genes, in determination of anticancer drug selectivity and toxicity.
However, there is a need to identify additional channels, or transporters, that are found in specific cancers, to enable the targeting of different cancers with anticancer agents that are substrates for those transporters.